Welcome to International Network for Natural Sciences | INNSpub

Paper Details

Research Paper | November 1, 2011

| Download

Extraction of β-galactosidase and β-glucosidase from the seeds of Tamarindus indica.

Shlini Purushothaman, Siddalinga Murthy KR.

Key Words:

Int. J. Biomol. & Biomed.1(3), 8-17, November 2011


IJBB 2011 [Generate Certificate]


The enzymes β–galactosidase and β–glucosidase were extracted from the tamarind seeds using different buffers at different pH. Highest activity was obtained with 10 mM sodium acetate buffer, pH 5.6 and 10 mM tris buffer, pH 7.4. The effect of NaCl and Triton X–100 at different concentrations on the extraction of the enzymes indicated 10 mM sodium acetate buffer, pH 5.6 containing 1 M NaCl as a better extractant of the enzyme. The enzyme assay was carried out using p–nitrophenyl–β–D–galactoside and p–nitrophenyl–β–D–glucoside as substrates. Highest enzyme activities were observed on 6th and 24th day of germination. The protein content gradually decreased upto 5th day of germination and suddenly increased on 6th day. However, on subsequent days of germination, the protein content greatly decreased upto 11th day. During the latter period of germination (18th day onwards) the content remained almost constant. The kinetic parameters varied for both β–galactosidase and β–glucosidase. The activity of β– galactosidase was show to have an optimal operating condition at pH 5.5 and a temperature of 500C. The thermostability of the enzyme was in the range of 400C – 700C with the pH stability in the range of 5.0 – 7.0. The Km and Vmax values for pNPGal were determined as 66μM and 2.27nmolesmin-1. In contrast the activity of β– glucosidase was shown to have an optimal operating condition at pH 5.0 and a temperature of 300C. The thermostability of the enzyme was in the range of 270C – 350C with the pH stability in the range of 4.0 – 7.0. The Km and Vmax values for pNPGlu were determined as 121μM and 5.26nmolesmin-1. The presented study is a preliminary work carried out for the standardization of protocols. The purification and characterization of β–galactosidase and β– glucosidase is under progress.


Copyright © 2011
By Authors and International Network for
Natural Sciences (INNSPUB)
This article is published under the terms of the Creative
Commons Attribution Liscense 4.0

Extraction of β-galactosidase and β-glucosidase from the seeds of Tamarindus indica.

Agrawal KM, Bahl OP. 1968. Glycosidases of Phaseolus vulgaris.II. isolation and general properties. Journal of Biological Chemistry. 243, 103-111.

Bhalla PL, Dalling MJ. 1984. Characteristics of a b-galactosidase associated with the stroma of chloroplasts prepared from mesophyll protoplasts of the primary leaf of wheat. Plant Physiol 76, 92–95.

Bonnin E, Lahaye M, Vigouroux J, Thibault JF. 1995. Preliminary characterization of a new exo-b-(1,4)-galactanase with transferase activity. Int J Biol Macromol 17, 345–351.

Buckeridge MS, Reid JS. 1994. Purification and properties of a novel b-galactosidase or exo-(134)-b-d-galactanase from the cotyledons of germinated Lupinus angustifolius L. seeds. Planta 192, 502–511.

Buckeridge MS, Rocha DC, Reid JSG, Dietrich SMC. 1992. Xyloglucan structure and post-germinative metabolism in seeds of Copaifera langsdorfii from savanna and forest populations. Physiol. Plant. 86, 1693-1702.

Carratu G, Colacino C, Conti S, Giannattasio M. 1985. Phytochem. 24, 1465.

Chengappa S, Jarman C, Fanutti C, Reid JSG. 1993. Xyloglucan oligosaccharide-specific a-D-xylosidase: molecular mode of action and cloning of a cDNA from germinated nasturtium (Tropaeolum majus L.) seeds. J. Cell. Biochem. (Suppl. 17A), 27.

Coronel RE. 1991. Tamarindus indica L. In Plant Resources of South East Asia, Wageningen, Pudoc. No.2. Edible fruits and nuts. (Eds.) Verheij, E.W.M. and Coronel, R.E., PROSEA Foundation, Bogor, Indonesia: 298-301.

David LS, Kenneth CG. 2000. A family of at least seven β-galactosidase genes is expressed during tomato fruit development. Plant Physiology, Vol.123, 1173-1183.

De Veau EJI, Gross KC, Huber DJ, Watada AE. 1993. Degradation and solubilization of pectin by b-galactosidases purified from avocado mesocarp. Physiol Plant 87, 279–285.

Dong HL, Sang-Gu K, Sang-Gon S, Jae KB. 2003. Purification and characterization of a b-galactosidases from Peach (Prunus persica). Mol. Cells. 15(1). 68-74.

Dumortier V, Brassart C, Bouquelet S. 1994. Purification and properties of a b-D-galactosidase from Bifidobacterium bifidum exhibiting a transgalactosylation reaction. Biotechnol. Appl. Biochem. 19, 341–354.

Edwards M, Bowman YJL, Dea ICM, Reid JSG. 1988. A b-galactosidase from nasturtium (Tropaeolum majus L.) cotyledons. Purification, properties and demonstration that xyloglucan is the natural substrate. J. Biol.Chem. 263, 4333–4337.

Edwards M, Dea ICM, Bulpin PV, Reid JSG. 1986. Purification and properties of a novel xyloglucan-specific endo-(1®4) -b-D-glucanase from germinated nasturtium seeds (Tropaeolum majus L.). J. Biol. Chem. 261, 9489–9494.

Edwards M, Dea ICM, Bulpin PV, Reid JSG. 1985. Xyloglucan (amyloid) mobilization in the cotyledons of Tropaeolum majus L.seeds following germination. Planta. 163, 133-140.

Erickson RP, Steers  E. Jr. 1970. Arch. Biochem. Biophys. 137, 339.

Esen A. 1993. Β-Glucosidases : Biochemistry and Molecular Biology, edited by A Esen, Washington, DC : American Chemical Society, p. 1-14.

Fanutti C, Gidley MJ, Reid JSG. 1991. A xyloglucanoligosaccharide-specific a-D-xylosidase or exo-oligoxyloglucan-a-xylohydrolase from germinated nasturtium (Tropaeolum majus L.) seeds. Planta, 184, 137–147.

Fanutti C, Gidley MJ, Reid JSG. 1993. Action of a pure xyloglucan endo-transglycosylase (formerly called xyloglucanspecific endo-(1®4)-b-D-glucanase from the cotyledons of germinated nasturtium seeds. Plant J. 3, 691–700.

Fanutti C, Gidley MJ, Reid JSG. 1996. Substrate subsite recognition of the xyloglucan endo-transglycosylase orxyloglucan-specific endo-(1®4)-b-D-glucanase from the cotyledons of germinated nasturtium (Tropaeolum majus L.) seeds. Planta, 200, 221–228.

Farkas V, Sulova Z, Stratilova E, Hanna R, Maclachlan G. 1992. Cleavage of xyloglucan by nasturtium seed xyloglucanase and transglycosylation to xyloglucan subunit oligosaccharides. Arch. Biochem. Biophys. 298, 365–370.

Giannakouros T, Karagiorgos A, Simos G. 1991. Expression of β-galactosidase multiple forms during barley (Hordeum vulgare) seed germination. Seperation and characterization of enzyme isoforms. Physiologia Plantarum. 82, 413-418.

Glasgow LR, Paulson JC, Hill RL. 1977. J.Biol. Chem. 252, 8615.

Hall BG. 1998. Determining the evolutionary potential of a gene. Mol Biol Evol 15, 1055–1061.

Harley S M, Beevers H. 1985. Characterisation and partial purification of three galactosidases from castor bean endosperm. Phytochemistry. 24, 1459-1464.

Hazel J Crombie, Sumant Chengappa, Amanda Hellyer and J S Grant Reid. 1998. A xyloglucan oligosaccharide-active, transglycosylating β-D-glucosidase from the cotyledons of nasturtium (Tropaeolum majus L) seedlings-purification, properties and characterization of a cDNA clone. The Plant Journal. 15(1) : 27-38.

Hirata H, Negoro S, Okada H. 1984. J. Bacteriol. 106, 9.

Huber RE, Kurz G, Wallenfels K. 1976. A quantitation of the factors which affect the hydrolase and transgalactosylase activities of b-galactosidase (E. coli) on lactose. Biochemistry 15, 1994–2001.

Kim YS, Lee EB Joo SH. 1991b. Arch. Pharm. Res. 14, 255.

Kim YS, Lee MY, Park YM. 1992. Kor. Biochem. J. 25, 171.

Li SC, Mazzotta MY, Chien SF, Li YT. 1975. J. Biol. Chem. 250. 6786.

Lo JT, Mukerji K, Awasthi YC, Hanada E, Suzuki K, Srivastava SK. 1979. Purification and properties of sphingolipid b-galactosidase from human placenta. J Biol Chem 254, 6710–6715.

Maley F, Trimble RB, Tarentino AL, Plummer TH. 1989. Characterization of glycoproteins and their associated oligosaccharides through the use of endoglycosidases. Anal Biochem 180, 195–204.

Masayuki S, Kiyoshi O, Yoichi T, Yohichi H, Shigeru Y. 1989. A β-Galactosidase fromm Radish (Raphanus sativus L.) Seeds. Plant Physiol. 90, 567-574.

Masayuki S, Atsushi I, Hajime I. 2000. Purification and characterization of a β-glucosidase from rye (Secale cereal L.) seedlings. Plant Science.155, 67-74.

Nagano H, Osmori M, Shoji Z, Kawaguchi T, Arai M. 1992. Biosci. Biotech. Biochem. 56, 674.

Nakao M, Harada M, Kodama Y, Nakayama T, Shibano Y, Amachi T. 1994. Purification and characterization of a thermostable b-galactosidase with high transgalactosylation activity from Saccharopolyspora rectivirgula. Appl. Microbiol. Biotechnol. 40, 657–663.

Onishi  N,  Yamashiro A,  Yokozeki  K.  1995. Production of galactooligosaccharide from lactose by Sterigmatomyces elviae CBS8119. Appl. Environ. Microbiol. 61, 4022–4025.

Papet MP, Delay D, Monsigny M, Delmotte F. 1992. Biochimie. 74, 53.

Raghothama KG, Lawton KA, Goldsbrough PB, Woodson WR. 1991. Characterization of an ethylene-regulated flower senescence-related gene from carnation. Plant Mol Biol 17, 61–71.

Reid JSG, Edwards M, Dea ICM. 1988. Enzymatic modification of natural seed gums. In Gums and Stabilisers for the Food Industry (Phillips, G.O.,Wedlock, D.J. and Williams,P.A., eds). Oxford: IRL Press, p. 391–398.

Reis D, Vian B, Darzens D, Roland JC. 1987. Sequential patterns of intramural digestion of galactoxyloglucan in tamarind seedlings. Planta. 170, 60-73.

Ross GS, Redgwell RJ, MacRae EA. 1993. Kiwifruit b-galactosidase: isolation and activity against specific fruit cell-wall polysaccharides. Planta 189, 499–506.

Shankaracharya NB. 1998. Tamarind – Chemistry, Technology and Uses – a critical appraisal. Journal of Food Technology, 35(3), 193-208.

Shigeta S, Kubota H, Tamura H, Oka S. 1983. J. Biochem 94, 1827.

Shlini P, Siddalinga MKR. 2011. Extraction of phenolics, proteins and antioxidant activity from defatted tamarind kernel powder. Asian J. Research Chem. 4(6), 936-941.

de Silva J, Jarman CD, Arrowsmith DA, Stronach MS, Chengappa S, Sidebottom C, Reid JSG. 1993. Molecular characterization of a xyloglucan-specific endo-(1®4)-b-Dglucanase (xyloglucan endo-transglycosylase) from nasturtium seeds. Plant J. 3, 701–711.

Sing-Chung Li, Jiahn-Wern Han, Kuan-Chung Chen, Ching-San Chen. 2001. Purification and characterization of isoforms of β-galactosidases in mung bean seedlings. Phytochemistry. 57, 349-359.

Steers E, Cuatrecasas P, Pollard H. 1971. J.Biol. Chem. 246, 196.

Stone BA, Clarke AE. 1992. Chemistry and Biology of (1-3)-β-D-Glucan. La Trobe University Press, Melbourne.

Tanaka  Y,  Kagamishi  A,  Kiuchi  A,  Shoji  Z. 1988. Agric. Biol. Chem. 52, 1301.

Yeong SK, Kyung SP, Jong GK. 1993. Purification and characterization of β-galactosidase from green onion. Korean Biochem J. 26(7), 602-608.

Yoon JH, Ajisaka K. 1996. The synthesis of galactopyranosly derivatives with b-galactosidases of different origins. Carbohydr Res 292, 153–163.


Style Switcher

Select Layout
Chose Color
Chose Pattren
Chose Background